ABSTRACT NUP98 rearrangements are observed in up to 10% of the childhood acute myeloid leukemia (AML) and are associated with poor prognosis. The unmet clinical needs and the lack of an effective targeted therapy to the NUP98-rearranged leukemias emphasize the need for novel regimens. Our central hypothesis is that NUP98-fusion oncoproteins harbor critical functional regions that are essential to their leukemogenetic potential. We also expect that NUP98-fusions will drive specific transcriptional networks, and provide potential vulnerabilities that can be exploited using CRISPR-mediated genome editing approach. We will test our hypothesis using multiple levels of CRISPR-mediated genetic screens covering genome-wide, target gene panel, and saturation protein scan. In collaboration with the Project 1 and Project 2 in this proposal, the Genome Editing Core will help provide critical insights into the molecular mechanisms by which NUP98- fusion oncoproteins lead to the development of leukemia. This work is innovative in that it will illuminate critical domains/motifs in NUP98-fusion oncoproteins, a particularly malignant disease that currently has no effective therapy. We expect that successful completion of this proposal will (1) establish a new genetic screen approach ?saturation CRISPR protein scan? for a sub-protein level functional domain discovery, and (2) yield novel mechanistic information about the functional regions in the NUP98-fusion oncoproteins. The impact of this research will be of significance because (1) it provides novel therapeutic opportunities against the difficult-to-treat NUP98-rearranged leukemias, and (2) it will help identify novel functional elements in fusion oncoproteins and their associated pathways for future pharmaceutical targeting.